Genetically engineered mesenchymal stem cells stably expressing green fluorescent protein

Mesenchymal stem cells [MSCs] are nonhematopoietic stromal cells that are capable of differentiating into and contribute to the regeneration of mesenchymal tissues. Human mesenchymal stem cells [liMSCs] are ideal targets in cell transplantation and tissue engineering. Enhanced green fluorescent protein [EGFP] has been an important reporter gene for gene therapy. The aim of this study was establishment of MSCs expressing GFP. MSCs were isolated and characterized by Immunophenotyping. The pEGFP-Nl plasmid was extracted from previously transformed Escherichia, coli cells and transfected into MSCs using FuGENE HD transfection reagent. Stable cells were established in the presence of geneticin. Expression of GFP was detected by RT-PCR, western blot analysis and immunoflorecent microscope. MSCs were successfully isolated and characterized. The MSCs transfected with the pEGFP-Nl plasmid expressed GFP both in mRNA and protein levels while cells transfected with empty vector did not. The results suggested that this engineered cell line will be used in the future studies and can easily be traced in vivo

Screening of soil and sheep faecal samples for predacious fungi: Isolation and characterization of the nematode-trapping fungus arthrobotrys oligospora

Over one-year period, 150 pasture soil samples and 138 sheep faecal samples, collected from different parts of Iran were screened for the presence of nematophagous fungi. The samples were cultured at 25°C on chloramphenicol-2% water agar [CHF-WA] plates in the presence of Haemonchus contortus third stage larvae [L3] and checked over a two-month period for characteristic conidia, conidiophores and hyphal traps of nematophagous fungi. Suspected nematophagous fungi were isolated by periodic transfer of the fungi on CHF-WA plates using the agar block method. Overall, 11 soil samples were found to harbour the nematode-trapping fungus Arthrobotrys from which 3 pure isolates were made and consequently identified as Arthrobotrys oligospora IRAN 877 C, IRAN 878 C and IRAN 879 C. Nematophagous fungi were not found in any tested sheep faecal samples. The predatory capacity of the isolates was tested against H. contortus infective larvae and then compared to reference strains A. oligospora CBS 111.37, A. oligospora CBS 251.82 and Duddingtonia flagrans CBS 583.91. The local strains of A. oligospora reduced the development of H. contortus L3 by 75-85%, whereas, the predatory capacity of reference A. oligospora and D. flagrans strains was measured in the range of 51-85% compared to the fungus free controls. Study of the effect of temperature on predatory activity of A. oligospora strains IRAN 877 C and CBS 111.37 revealed a reduction of more than 95% in infective larvae of H. contortus at temperature levels between 15 to 25°C. This reduction was significantly decreased to 30% and 50% at 10°C and 30°C, respectively. The nematode-trapping fungus A. oligospora is reported from soil of Iran for the first time and its potential role in biocontrol of gastrointestinal nematodes of ruminants is discussed